The overall objective of this competing R01 renewal is to exploit a newly discovered, general mechanism of attachment-triggered self-assembly of hydrophobic drugs to create thermally responsive nanoparticles for the treatment of primary and metastatic cancer. In the previous funding cycle, we discovered that the attachment of multiple copies of hydrophobic chemotherapeutics such as doxorubicin (Dox) and paclitaxel (PTX) at the chain end of a recombinant chimeric polypeptide (CP) resulted in the spontaneous self-assembly of the CP- drug conjugate into near monodisperse, highly soluble nanoparticles. CP-Dox nanoparticles showed dramatic efficacy in curing sub-cutaneous (s.c.) murine colon carcinoma (>90%) in mice and some efficacy in treating metastatic breast cancer in an orthotopic model. These results are highly encouraging but suggest that passive targeting alone of a single drug is unlikely to be therapeutically sufficient. We hence propose to build upon the success of this approach to greatly improve the efficacy of CP-drug nanoparticles by exploiting thermal targeting to co-deliver two complementary chemotherapeutics -Dox and paclitaxel (PTX)- by CP-drug nanoparticles to treat primary and metastatic breast cancer in an orthotopic disease model. To impart thermal targeting capabilities to P-Dox nanoparticles, we have re-engineered the composition and chain length of the CPs to undergo selective and reversible aggregation in the vasculature of tumors that are externally heated to 420C. We propose to apply multiple cycles of mild hyperthermia to pump the CP-drug nanoparticles out of the tumor vasculature and into the tumor interstitium. The proposed research will test the hypotheses that: (1) combination chemotherapy is superior to monotherapy; (2) thermal targeting will provide increased tissue-level accumulation and better control of primary tumor that will limit dissemination of metastases from the primary tumor. This research is innovative because, to our knowledge, a rationally engineered drug-nanoparticle system that can be thermally targeted to solid tumors has not been demonstrated for a single drug, let alone multiple types of chemotherapeutics. The impact of this research will be the development of a targeted nanoparticle drug delivery platform that will demonstrate therapeutic efficacy in a preclinical orthotopic tumor model for primary and metastatic disease with two different chemotherapeutics, and will position this technology to graduate to a clinical trial at the end of this funding period.